α-d-Glucopyranosiduronic acid, (3β,20β)-20-carboxy-11-oxo-30-norolean-12-en-3-yl 2-O-β-d-glucopyranuronosyl-, dipotassium salt

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

α-d-Glucopyranosiduronic acid, (3β,20β)-20-carboxy-11-oxo-30-norolean-12-en-3-yl 2-O-β-d-glucopyranuronosyl-, dipotassium salt

EC Number:

272-296-1

EC Name:

α-d-Glucopyranosiduronic acid, (3β,20β)-20-carboxy-11-oxo-30-norolean-12-en-3-yl 2-O-β-d-glucopyranuronosyl-, dipotassium salt

Cas Number:

68797-35-3

Molecular formula:

C42H62O16.2K

IUPAC Name:

α-d-Glucopyranosiduronic acid, (3β,20β)-20-carboxy-11-oxo-30-norolean-12-en-3-yl 2-O-β-d-glucopyranuronosyl-, dipotassium salt

Specific details on test material used for the study:

Batch number: 3194/001/A17
Appearance: White to yellowish Powder
Purity: 95% (102.7% according to the Certificate of Analysis)
Expiry date: 11 July 2019
Storage conditions: Controlled room temperature (15-25°C, ≤70% relative humidity), protected from light and humidity (store in a tightly closed container)

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Analytical measurements were performed from the control and at the applied test concentration level at the beginning and at the end of the experiment.

Test solutions

Details on test solutions:

Reconstituted water (ISO medium, according to OECD 202) was used as dilution water for both the range finding and definitive tests. The same composition of reconstituted water was used for the tests and for breeding the test animals.
A stock solution with a nominal concentration of 100.0 mg/L was prepared with direct addition of the test item, mixed into the test medium (ISO Medium) using ultrasonic bath for approximately 2 minutes. As a Limit test was carried out, further dilution of stock solution was not performed.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

Source: István Szent University, 2100 Gödöllő, Páter Károly u. 1, Hungary
Breeding: The Daphnia are bred in Ecotoxicological Laboratory of Citoxlab Hungary Ltd. The health of the stock animals is continuously monitored by visual daily checking. Abnormal behaviour or significant decrease of population is recorded.
Justification of strain: Daphnia magna is the standard species of the acute immobilisation test.
Number of animals: There were 20 animals in test group and control group respectively, divided into 4 replicates (5 animals / replicate)
Age of the animals: They were less than 24 h old at the beginning of the test
Acclimatization: There was no acclimatization because the water used was similar to the culture water.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Test conditions

Hardness:

The reconstituted water (ISO medium) had a total hardness of 248 mg/L (as CaCO3).

Test temperature:

The water temperature was measured at the start of the experiment and in 24-hour intervals thereafter in each test vessel. The test temperature was in the range of 20.4 – 20.7°C measured in the test vessels.
The additionally measured temperature in the climate chamber was between 20.0 – 20.9°C.

pH:

The pH of the test solution was not adjusted and not varied by more than 1.5 units in any one test. The pH was measured at the start and at the end of the experiment in each test vessel and was in the range of 7.22 – 7.35.

Dissolved oxygen:

The dissolved oxygen concentration was measured in each test vessel at the start and at the end of the test and was in the range of 8.2 – 8.7 mg/L.

Nominal and measured concentrations:

As the measured concentration deviated not more than 20 per cent from the nominal, biological results are based on the nominal test item concentration.

Details on test conditions:

- Twenty animals, divided into four groups (glass beaker) of five animals each were used at the test concentration and for the control.
- Because no toxic response was observed during the preliminary range-finding test, a Limit Test was carried out using only one test concentration (100.0 mg/L) and one control group in the definitive test.
- Light: The test was carried out in 16-hour light and 8-hour dark cycle.
- Observations: The immobility or mortality of the Daphnia was determined by visual observation 24 and 48 hours after the start of the test. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker are considered to be immobile.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 2

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 3

Key result

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 4

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

ca. 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 5

Key result

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Any other information on results incl. tables

* ANALYTICAL CONCENTRATIONS

The test concentration was analytically determined at the start and at the end of the experiment. Measured concentration was 101.91 mg/L at the start and 104.87 mg/L at the end of the experiment.

* VALIDITY OF THE TEST

There were no immobilized animal in the control group and the dissolved oxygen concentration at the end of the test in control and test vessels was more than 3 mg/L.

All validity criteria were within acceptable limits and therefore the study can be considered as valid.

* IMMOBILISATION

The number of immobilised animals and the percentage of immobility were determined at the 24^thand 48^thhour (see the table 1 below).

In addition to immobility, no abnormal behaviour or appearance of test animals was detected.

Table 1: Number and percentage of immobilised animals

Test Group	Number of treated animals	Number of immobilised animals
		24 hours		48 hours
		number	percent	number	percent
Control	20	0	0	0	0
100.0 mg/L	20	0	0	0	0

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Acute toxicity of Dipotassium Glycyrrhizate on Daphnia magna was assessed in an Acute immobilisation test, over an exposure period of 48 hours in a static test system.
Under the conditions of this Daphnia magna acute immobilisation study the observed endpoints for the effect of Dipotassium Glycyrrhizate were the followings:
- The 24h and 48h EC50 value > 100.0 mg/L (nominal)
- The 48h EC100 value > 100.0 mg/L (nominal)
- The 48h No-Observed Effect Concentration (NOEC) = 100.0 mg/L (nominal)
- The 48h Lowest Observed Effect Concentration (LOEC) > 100.0 mg/L (nominal)